Phylogenetic affiliations of marine fungi detected with pyrosequencing and ion semiconductor targeted amplicon sequencing
K. T. PICARD (1), R. F. Stern (2), F. Lutzoni (1). (1) Duke University, Durham, NC, U.S.A.; (2) Sir Alistair Hardy Foundation for Ocean Science, Plymouth, United Kingdom
The systematic relationships of zoosporic fungi (Blastocladiomycota, Chytridiomycota, Cryptomycota, Neocallimastigomycota, and <i>Olpidium</i>) are known almost exclusively from taxa isolated in culturing surveys of terrestrial and freshwater habitats. Recent culture-independent molecular cloning studies of extreme marine habitats, such as deep-sea hydrothermal vents and methane seeps, have revealed novel fungal phylotypes belonging to the zoosporic lineages. However, as environmental cloning studies frequently fail to capture all of the diversity within a given sample, our understanding of both the breadth of zoosporic fungal diversity and the phylogenetic placement of marine taxa remains poor. Short (200-600bp) 454 reads from nrSSU were generated from water samples from the English Channel using general eukaryotic primers. From marine and estuarine sediments from coastal North Carolina, 330bp nrLSU reads were generated on the Ion Torrent sequencing platform using fungal-affinity primers. Using maximum likelihood and Bayesian methods, the phylogenetic affinities of marine fungi, represented by these nuclear ribosomal RNA-coding sequences, were inferred. Extensive kingdom-wide reference alignments and multiple backbone constraint trees for both loci were used in our analyses to ensure accurate placement of short reads for OTU estimation. These results will be compared to inferred phylogenies based on long (Sanger) reads (1.2 kb) from cloned amplicon fragments of the nrLSU.
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